Chitosan nanoparticles as a targeted delivery system for anti-fibrotic microRNAs for oral submucosal fibrosis treatment

Abstract

Oral submucous fibrosis (OSF) is characterized by excessive extracellular matrix (ECM) deposition. Dysregulation of microRNAs (miRs) is involved in the progression of OSF, and miR manipulation could be a promising therapeutic approach. Nanoformulation can protect exogenous miRs against nuclease degradation and enhance cell retention. Accordingly, chitosan (CS), which possesses an anti-fibrotic capacity, is proposed to encapsulate miRs as nanoparticles (NPs) for treating OSF. miR-negative control (miR-NC)/CS NPs were fabricated by an ionic gelation method and characterized. Human oral submucosal fibroblasts were first subjected to arecoline stimulation to induce myofibroblast differentiation and were then transfected with a miR-145 inhibitor or miR-424 inhibitor using CS NPs. For CS NPs loaded with miR-NC, the particle size was 121.9 ± 0.1 nm with a polydispersity index of 0.162 ± 0.004 and zeta potential of + 22.4 ± 0.5 mV. Transfection of these two miRs downregulated mRNA levels of transforming growth factor beta 1, actin alpha 2 smooth muscle, collagen type I alpha 1 chain (COL1A1), COL3A1, COL4A1, matrix metalloproteinase 2, tissue inhibitor of metalloproteinase 2, and zinc finger E-box binding homeobox 1 in myofibroblasts. A Western blot analysis revealed that miR/CS NP transfection decreased alpha-smooth muscle actin and type 1 collagen protein products. Furthermore, the wound closure ability of stimulated cells was inhibited upon transfection. In conclusion. CS NPs are a good delivery vehicle for miR transfection. Transfection of a miR-145 inhibitor and miR-424 inhibitor inhibited the TGF-β signaling pathway and decreased ECM component production, and could thus be a promising treatment for OSF.